Simultaneous determination of methylenetetrahydrofolate reductase C677T and factor V G1691A genotypes by mutagenically separated PCR and multiple-injection capillary electrophoresis.
The C677T mutation of the methylenetetrahydrofolate reductase gene and the G1691A (Leiden) mutation of the factor V gene are established risk factors for thromboembolic disease. We here present an assay for the simultaneous genotyping of these common genetic variants. The assay involves a strategy based on multiplex mutagenically separated PCR performed in a single tube containing six primers. Separation of the resulting four PCR products (197, 207, 233, and 246 bp) was performed by capillary electrophoresis coupled with laser-induced fluorescence detection. The time for the automated electrophoresis was reduced to 2.5 min per sample by performing the capillary electrophoresis analysis in a multiple-injection mode.
Ulvik
A
A
Department of Pharmacology, University of Bergen, Norway.
Ren
J
J
Refsum
H
H
Ueland
P M
PM
D001331:Automation; D004252:DNA Mutational Analysis / Q000379:methods*; D017931:DNA Primers / Q000737:chemistry; D019075:Electrophoresis, Capillary; D005165:Factor V / Q000235:genetics*; D005453:Fluorescence; D005838:Genotype; D006710:Homocysteine / Q000097:blood; D007834:Lasers; D042965:Methylenetetrahydrofolate Reductase (NADPH2); D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012307:Risk Factors; D013927:Thrombosis / Q000235:genetics
D017931:DNA Primers; C095381:factor V Leiden; D006710:Homocysteine; D005165:Factor V; D000587:Oxidoreductases Acting on CH-NH Group Donors; D042965:Methylenetetrahydrofolate Reductase (NADPH2)